Fourteen essential oils (EOs) from selected live germplasm of medicinal plants have been tested for their antitrypanosomal and cytotoxic activity. These plants have been domesticated and maintained under experimental cultivation. Their EOs were tested on epimastigote forms of Trypanosoma cruzi strain Y and human lung fibroblasts LC5 cell line, along with the major components of the active oils, both separately and in binary combinations. Mentha rotundifolia, Thymus zygis, T. vulgaris and Hyssopus officinalis were the most active EOs against T. cruzi. Among the main components of these EOs (1-8-cineole, thymol, p-cymene, piperitenone oxide, β-pinene, γ-terpinene, carvacrol and linalool), the most active against the parasite and less toxic to human cells was thymol. In general, the activity of the main components did not exceed that of their origin EO, and the study of the activity of these compounds in combination indicates the existence of antagonistic and synergistic effects depending on the concentration tested.